Improving cognitive impairment by Tongxinluo via inhibiting expression of beta-secretase 1/beta-amyloid peptide in experimental vascular dementia

BACKGROUND： Tongxinluo has been clinically proven to be effective in improving memory and cognitive function in patients with post-stroke vascular dementia. Is the mechanism related to the deposition of beta-amyloid peptide （A β ） in hippocampus？ OBJECTIVE： To observe the effect of Tongxinluo on cognitive impairment in a mouse model with vascular dementia and the changes of A β deposition and β -secretase 1 （BACE1） expression. DESIGN： Randomized controlled study. SETTING： State Key Laboratory of Pharmaceutical Biotechnology of Nanjing University and Affiliated Drum Tower Hospital of Nanjing University Medical School. MATERIALS： The experiment was carried out in the State Key Laboratory of Pharmaceutical Biotechnology of Nanjing University and Affiliated Drum Tower Hospital of Nanjing University Medical School from March 2006 to January 2007. A total of 36 healthy Kunming mice, 18 of each gender, were chosen. The study was conducted in accordance with the National Regulations of Experimental Animal Administration, and all animal experiments were approved by the Committee of Experimental Animal Administration of Nanjing University. Tongxinluo was provided by Shijiazhuang Yiling Pharmaceutical Co., Ltd. METHODS： All mice were randomly divided into 6 groups, including naive control （n=6）, sham-operated control （n=6） and experimental groups treated with different doses of Tongxinluo （0.2, 0.4, and 0.6 g/kg/d; n=6 for each group） or vehicle （n=6）. Five groups were subjected to bilateral common carotid arteries （2-VO） occlusion to produce a vascular dementia model （no occlusion was performed in sham-operated group）. The mice in the Tongxinluo treatment groups were intragastricly administered daily with a Tongxinluo suspension （40 g/L in distilled water） at doses of 0.2, 0.4 or 0.6 g/kg/d from day 1 to day 30 post-surgery. The animals in vehicle, sham-operated and naive groups were administered an equal volume of distilled water. MAIN OUTCOME MEASURES： ①Escape latency time determined in all groups of mice before and after 2-VO occlusion by Morris water maze. ②Changes in BACEI mRNA expression in the hippocampi of mice among the six groups by RT-PCR assay, and BACEI and A β protein expression in the hippocampi of mice by Western blot. RESULTS： All 36 mice were involved in the final analysis.① No difference was detected in escape latency time to a hidden platform among all groups in water maze test before surgery （P 〉 0.05） At 30 days after 2-VO occlusion, the vehicle animals exhibited a significantly longer latency in finding the hidden platform compared to that of sham-operated and naive animals （P 〈 0.01）. The prolonged escape latency was significantly reduced by oral administration of 0.4 g or 0.6 kg/day （P 〈 0.01, P 〈 0.05）. BACEI mRNA and protein expression in vehicle animals were much higher than in sham-operated and naive animals （P 〈 0.01）. The ischemia-induced increases in BACE1 mRNA and protein level were attenuated by all three doses of Tongxinluo treatment （P 〈 0.01）, and the 0.4 g/kg/d treatment was the most effective. A β protein expression in vehicle animals after 2-VO occlusion were much higher than in sham-operated and naive animals （P 〈 0.01）. 2-VO occlusion-induced A β generation was significantly attenuated by all doses of Tongxinluo treatment, with the most effective dose being 0.4 g/kg/d （P 〈 0.01）. CONCLUSION： BACE1 mRNA levels and protein levels of BACEI and A β are reduced in the hippocampi of vascular dementia model mice by all three doses of Tongxinluo treatment, with the most effective dose being 0.4 g/kg/d. The results suggest that inhibition of post-ischemia BACEI expression and A β generation in brain might underlie Tongxinluo＇s effects in improving cognitive impairment.

Neuroprotective effect of the traditional Chinese herbal formula Tongxinluo: a PET imaging study in rats

Tongxinluo has been widely used in China for the treatment of acute stroke and for neuroprotection. However, there are few positron emission tomography （PET） studies on the neuroprotective effect of Tongxinluo on cerebral ischemia/reperfusion in small animals. In the present study, Tongxinluo superfine powder suspension or its vehicle was administered intragastrically to rats for 5 successive days before middle cerebral artery occlusion, ^18Ffluorodeoxyglucose （FDG） small animal PET imaging showed that at 1 and 2 weeks after cerebral ischemia/reperfusion, glucose metabolism in the ischemic area was greater in rats that had received Tongxinluo than in those that had received the vehicle. Nissl staining showed that 2 weeks after cerebral ischemia/reperfusion, there was less neuronal loss in the prefrontal cortex in Tongxinluo-treated rats than in controls. In addition, Tongxinluo-treated animals showed better neurologic function and lower cerebral infarct volume than rats that received the vehicle. These findings suggest that Tongxinluo exhibits neuroprotective effects in cerebral ischemia/reperfusion injury and demonstrates that ^18F-FDG small animal PET imaging is a useful tool with which to study the molecular pharmacology of traditional Chinese medicine.

Intervention of Tongxinluo Capsule (通心络胶囊) against Vascular Lesion of Atherosclerosis and Its Effect on Lectin-like Oxidized Low Density Lipoprotein Receptor-1 Expression in Rabbits

Objective.- To investigate the prevention by Tongxinluo capsule （通心络胶囊, TXL） of vascular lesions and its effect on the levels of protein and gene expression of lectin-like oxidized low-density lipoprotein receptor-1 （LOX-1） of vascular wall in rabbits with atherosclerosis （AS）, and to explore its possible mechanism against AS. Methods： AS models were established by feeding New Zealand white rabbits with high-cholesterol diet, and 24 immature rabbits were randomly divided into the control group, model group and treated group （treated with TXL capsule）. The indexes of total cholesterol （TO） and low density lipoprotein （LDL） levels were measured at the 16th week. The intima thickness and the plaque area of abdominal aorta were quantitatively analyzed by pathological morphological analysis, the expression of macrophage and smooth muscle cell （SMC） in intima were detected by immunohistochemical method and histologic segments were stained by Hematoxilin-Eosin （HE） to identify the degree of atherosclerotic lesion in the model group and the prevention by TXL. The LOX-1 gene and protein expression in abdominal aorta was detected by semi-quantitative RT-PCR and immunohistochemistry, respectively. Results： In the model group, the levels of TC and LDL were significantly elevated, aortic intima thickened extensively, the intima area enhanced, and macrophages expression increased; the levels of LOX-1 gene and protein expression was up-regulated in endothelium and neo-intima of the abdominal aorta. The treatment with TXL reduced blood lipids, attenuated arterial intimal proliferation, markedly inhibited the expression of macrophage and excessively expressed the level of LOX-1. Conclusion：TXL has an inhibitory effect on blood lipids, and it can prevent the occurrence of vascular lesion and cure its development, and its protection against AS was possibly associated with a crucial endothelial protective action through lowering the expression of LOX-1 in vascular walls.

Effects of Digoxin and Tongxinluo capsule combination treatment on oxidative stress, cytokines and vascular endothelial function in patients with chronic heart failure

Objective:To explore the effect of digoxin and Tongxin capsule combined therapy on oxidative stress, cytokines and vascular endothelial function in patients with chronic heart failure, and provide help for clinical treatment of patients with chronic heart failure.Methods:95 cases of chronic heart failure in our hospital were randomly divided into observation group (47 cases) and control group (48 cases). Control group patients were given basic treatment, and observation group received combination therapy of digoxin and Tongxinluo capsule, to detect and to investigate the changes of oxidative stress, cytokines and vascular endothelial function in two groups of patients before and after treatment.Results:There was no significant difference in oxidative stress, cytokines and vascular endothelial function between the two groups of chronic heart failure patients before treatment (P>0.05). Compared with before treatment, the malondialdehyde (MDA) and cytokines [tumor necrosis factor-α (TNF-α), C-reactive protein (CRP), interleukin-6 (IL-6) and brain natriuretic peptide (BNP)] in two groups of patients with chronic heart failure after treatment decreased significantly (P<0.05), while indexes related to endothelial function [hyperemic brachial artery diameter after reactive hyperemia, brachial artery diameter change rate, brachial artery endothelial dependent diastolic function (FMD), and brachial artery endothelium dependent diastolic function (NMD)] and related indexes of oxidative stress [glutathione peroxidase (GSH-Px), superoxide dismutase (SOD), and catalase (CAT)] were significantly increased (P<0.05). The related indexes and oxidative stress indexes (GSH-Px, CAT and SOD) of observation group after the combined treatment in patients with endothelial function were significantly higher than the control group after treatment (P<0.05). After treatment, cytokines and MDA levels were significantly lower than the control group (P<0.05).Conclusions:Tongxinluo capsule combined with digoxin treatment significantly improved the oxidative stress, cytokines and vascular endothelial function levels in patients with chronic heart failure, and has important clinical significance for the treatment of patients with chronic heart failure.

Clinical study on the effect of Tongxinluo combined with trimetazidine on cardiac function in patients with acute ST-segment elevation myocardial infarction after percutaneous coronary intervention

Objective:To investigate the clinical effect of Tongxinluo combined with trimetazidine on cardiac function in patients with acute ST-segment elevation myocardial infarction after percutaneous coronary intervention.Method: From March 2014 to September 2016, we selected 190 patients with ST-segment elevation myocardial infarction with percutaneous coronary intervention, according to the admission time is divided into observation group and control group, the control group was treated with conventional therapy (aspirin, isosorbide dinitrate, metoprolol tartrate, clopidogrel sulfate, captopril, atorvastatin calcium and diuretics) and trimetazidine, observation group in the control group based on Tongxinluo combined treatment, each group of 95 cases, and hs-CRP, aldosterone, NT-proBNP, TNF-α, IL-6, and cardiac function (LVEDV,LVESV,LVEF,SV) were compared.Result: The Hs-CRP in the observation group was significantly lower than that in the control group;The aldosterone in the observation group was significantly lower than that in the control group;The levels of NT-proBNP, TNF-α and IL-6 in the observation group were significantly lower than those in the control group;LVVEV and LVESV were significantly lower in the observation group than in the control group, LVEF and SV were significantly higher than those in the control group.Conclusion:Tongxinluo combined with trimetazidine in patients with acute ST-segment elevation myocardial infarction after percutaneous coronary intervention in patients with clinical effect is better, stable plaque, effectively improve microcirculation and cardiac function, recommended a wide range of clinical application.

No-reflow protection and long-term efficacy for acute myocardial infarction with Tongxinluo： a randomized double-blind placebo- controlled multicenter clinical trial （ENLEAT Trial）

Background No-reflow after emergency percutaneous coronary intervention （PCI） for acute ST segment elevation myocardial infarction （STEMI） is related to the severe prognosis. The aim of this study was to evaluate the efficacy of Tongxinluo, a traditional Chinese medicine, on no-reflow and the infarction area after emergency PCI for STEMI.Methods A total of 219 patients （female 31, 14%） undergoing emergency PCI for STEMI from nine clinical centers were consecutively enrolled in this randomized, double-blind, placebo-controlled, multicenter clinical trial from January 2007 to May 2009. All patients were randomly divided into Tongxinluo group （n=108） and control group （n=111）, given Tongxinluo or placebo in loading dose 2.08 g respectively before emergency PCI with asprin 300 mg and clopidogrel 300 mg together, then 1.04 g three times daily for six months after PCI. The ST segment elevation was recorded by electrocardiogram at hospitalization and 1, 2, 6, 12, 24 hours after coronary balloon dilation to evaluate the myocardial no-flow; myocardial perfusion scores of 17 segments were evaluated on day 7 and day 180 after STEMI with static single-photon emission computed tomography （SPECT） to determine the infarct area.Results There was no statistical significance in sex, age, past history, chest pain, onset-to-reperfusion time, Killip classification, TIMI flow grade just before and after PCI, either in the medication treatment during the follow up such as statin, β-blocker, angiotensin converting enzyme inhibitor （ACEI） or angiotensin receptor blocker （ARB） between two groups. There was significant ST segment restoration in Tongxinluo group compared to the control group at 6 hours （（-0.22±0.18） mV vs. （-0.18±0.16） mV, P=0.0394）, 12 hours （（-0.24 ± 0.18） mV vs. （-0.18±0.15） mV, P=0.0158） and 24 hours （（-0.27±0.16） mV vs. （-0.20±0.16） mV, P=0.0021） reperfusion; and the incidence of myocardial no-reflow was also reduced significantly at 24-hour reperfusion （34.3% vs. 54.1%, P=0.0031）. The myocardial perfusion scores of 17 segments evaluated by static SPECT was improved significantly on day 7 and day 180 after STEMI in Tongxinluo group compared to the control group （0.61±0.40 vs. 0.76±0.42, P=0.0109 and 0.51 ±0.42 vs. 0.66±0.43, P=0.0115, respectively）.There was no significant difference in severe adverse events between two groups.Conclusion Tongxinluo as a kind of traditional Chinese medicine could reduce myocardial no-reflow and infarction area significantly after emergency PCl for STEMI with conventional medicine therapy.

Effects of Tongxinluo-facilitated cellular cardiomyoplasty with autologous bone marrow-mesenchymal stem cells on postinfarct swine hearts

Background Treatment of ischemic heart disease remains an important challenge, though there have been enormous progresses in cardiovascular therapeutics. This study was conducted to evaluate whether Tongxinluo （TXL） treatment around the transplantation of mesenchymal stem cells （MSCs） can improve survival and subsequent activities of implanted cells in swine hearts with acute myocardial infarction （AMI） and reperfusion. Methods Twenty-eight Chinese mini-pigs were divided into four groups including a control group （n=7）; group 2, administration of low-close TXL alone from the 3rd day prior to AMI to the 4th day post transplantation （n=-7）; group 3, MSCs alone （n=-7） and group 4, TXL ＋ MSCs （n=7）. AMI models were made by occlusion of the left anterior descending coronary artery for 90 minutes. Autologous bone marrow-MSCs （3×10^7 cells/animal） were then injected into the post-infarct myocardium immediately after AMI and reperfusion. The survival and differentiation of implanted cells in vivo were detected by immunofluorescent analysis. The data of cardiac function were obtained at baseline （1 week after transplantation） and endpoint （6 weeks after transplantation） by single photon emission computed tomography （SPECT） and magnetic resonance imaging （MRI）. Apoptosis was detected by TUNEL assay and the oxidative stress level was investigated in the post-infarct myocardium at endpoint. Results At endpoint, there was less fibrosis and inflammatory cell infiltration with more surviving myocardium in group 4 than in the control group. In group 4 the survival and differentiation of implanted MSCs were significantly improved more than that seen in group 3 alone （P〈0.0001）; the capillary density was also significantly greater than in the control group, group 2 or 3 both in the infarcted zone （P〈0.0001） and the peri-infarct zone （P〈0.0001）. MRI showed that parameters at baseline were not significantly different between the 4 groups. At endpoint, regional wall thickening and the left ventricular ejection fraction were increased while the left ventricular mass index, dyskinetic segments and infarcted size were decreased only in group 4 compared with control group （P〈0.0001）. SPECT showed that the area of perfusion defect was significantly decreased at endpoint only in group 4 compared with control group （P〈0.0001）. TUNEL assay indicated that TXL administration significantly decreased cell apoptosis in peri-infarct myocardium in groups 2 and 4. Furthermore, superoxide dismutase （SOD） significantly increased and malondialdehyde （MDA） decreased in groups 2 and 4 by the administration of TXL. Conclusions Our study demonstrates the following： （1） immediate intramyocardial injection of MSCs after AMI and reperfusion resulted in limited survival and differentiation potential of implanted cells in vivo, thus being incapable of beneficially affecting post-hearts; （2） TXL-facilitation resulted in a significant survival and differentiation potential of implanted cells in vivo via inhibition of apoptosis and oxidative stress, accompanied by significant benefits in cardiac function.

Tongxinluo Capsule(通心络胶囊)for Cardiac Syndrome X：A Systematic Review and Meta-Analysis

Objective： To evaluate the efficacy and safety of Tongxinluo Capsule（通心络胶囊, TXL） for patients with cardiac syndrome X（CSX）. Methods： Randomized controlled trials（RCTs） regarding TXL in the treatment of CSX were searched in Chinese Biomedicine Literature Database, China National Knowledge Infrastructure, Chinese Scientific Journal Database, Wanfang Database, Pub Med, EMBASE, Cochrane Central Register of Controlled Trial, websites of the Chinese and International Clinical Trial Registry platform up to June 30, 2015. The intervention was either TXL alone or TXL combined with conventional treatment, while the control intervention was conventional treatment with or without placebo. Data extraction, methodological quality assessment and data analyses were performed according to the Cochrane criteria. The primary outcome was a composite event of death, acute myocardial infarction（AMI）, angina requiring hospitalization, revascularization, and heart failure. The secondary outcome measures were angina symptom improvement, electrocardiograph（ECG） improvement, and serum endothelin-1（ET-1） level. The adverse events were also recorded. RevMan 5.3 software was applied for data analyses. Results： Twelve RCTs（696 patients） were included. Compared with conventional treatment, the addition of TXL to conventional treatment showed some benefits on relieving angina symptoms [risk ratio（RR）： 1.46, 95% confidence interval（CI）（1.25, 1.71）, P〈0.01], and improving ECG [RR： 1.45, 95% CI（1.21, 1.74）, P〈0.01]. The pooled result did not support a benefit of TXL on reducing the incidence of primary outcome [RR： 0.20, 95% CI（0.02, 1.61）, P=0.13]. In addition, TXL decreased serum ET-1 concentration of CSX patients [standardized mean number： –1.63, 95% CI（–2.29, –0.96）, P〈0.01]. No serious adverse events were reported. Conclusions： TXL documents potential benefits on attenuating angina symptoms, improving ECG and decreasing serum ET-1 level for CSX patients. However, more rigorous RCTs with high quality are needed to confirm its efficacy and safety.

Tongxinluo Reverses the Hypoxia-suppressed Claudin-9 in Cardiac Microvascular Endothelial Cells

Background： Claudin-5, claudin-9, and claudin-11 are expressed in endothelial cells to constitute tight junctions, and their deficiency may lead to hyperpermeability, which is the initiating process and pathological basis of cardiovascular disease.Although tongxinluo （TXL） has satisfactory antianginal effects, whether and how it modulates claudin-5, claudin-9, and claudin-1 1 in hypoxia-stimulated human cardiac microvascular endothelial cells （HCMECs） have not been reported.Methods： In this study, HCMECs were stimulated with CoCl2 to mimic hypoxia and treated with TXL.First, the messenger RNA （mRNA） expression of claudin-5, claudin-9, and claudin-l 1 was confirmed.Then, the protein content and distribution of claudin-9, as well as cell morphological changes were evaluated after TXL treatment.Furthermore, the distribution and content histone H3K9 acetylation （H3K9ac） in the claudin-9 gene promoter, which guarantees transcriptional activation, were examined to explore the underlying mechanism, by which TXL up-regulates claudin-9 in hypoxia-stimulated HCMECs.Results： We found that hypoxia-suppressed claudin-9 gene expression in HCMECs （F=7.244;P =0.011） and the hypoxia-suppressed claudin-9 could be reversed by TXL （F=61.911;P =0.000）, which was verified by its protein content changes （F=29.142;P =0.000）.Moreover, high-dose TXL promoted the cytomembrane localization of claudin-9 in hypoxia-stimulated HCMECs, with attenuation of cell injury.Furthermore, high-dose TXL elevated the hypoxia-inhibited H3K9ac in the claudin-9 gene promoter （F=37.766;P =0.000）, activating claudin-9 transcription.Conclusions： The results manifested that TXL reversed the hypoxia-suppressed claudin-9 by elevating H3K9ac in its gene promoter, playing protective roles in HCMECs.

Tongxinluo Inhibits Cyclooxygenase-2, Inducible Nitric Oxide Synthase, Hypoxia-inducible Factor-2α/Vascular Endothelial Growth Factor to Antagonize Injury in Hypoxia-stimulated Cardiac Microvascular Endothelial Cells

Background： Endothelial dysflinction is considered as the initiating process and pathological basis of cardiovascnlar disease. Cyclooxygenase-2 （COX-2） and prostacyclin synthase （PGIS）, inducible nitric oxide synthase （iNOS） and endothelial NOS （eNOS） are key enzymes with opposing actions in inflammation and oxidative stress, which are believed to be the major driver of endothelial dysfunction. And in hypoxia （Hx）, Hx-inducible factor （HIF）-1α and HIF-2α are predominantly induced to activate vascular endothelial growth factor （VEGF）, restllting in abnormal proliferation. Whether and how Tongxinluo （TXL） modulates COX-2, PGIS, iNOS, eNOS, HIF-1α, HIF-2α, and VEGF in Hx-stimulated human cardiac microvascular endothelial cells （HCM ECs） have not been clarified. Methods： HCMEC were treated with CoCl2 to mimic Hx and the mRNA expressions of COX-2, PGIS, iNOS, eNOS, HIF-1α, HIF-2α. and VEGF were first confirmed, and then their mRNA expression and protein content as well as the cell pathological alterations were evaluated for TXL treatment with different concentrations, In addition, the effector molecular of inflammation prostaglandin E2 （PGE2） and the oxidative marker nitrotyrosine （NT） was adopted to reflect HCMEC in.jury. Results： Hx could induce time-dependent increase of COX-2, iNOS, HIF-2α, and VEGF in HCMEC. Based on the Hx-induced increase, TXL could mainly decrease COX-2, iNOS, HIF-2α, and VEGF in a concentration-dependent manner, with limited effect on the increase of PGIS and eNOS. Their protein contents verified the mRNA expression changes, which was consistent with the cell morphological alterations. Furthermore, high dose TXL could inhibit the Hx-induced increase of PG E, and NT contents, attenuating the inflammatory and oxidative injury. Conclusions： TXL could inhibit inflammation-related COX-2, oxidative stress-related iNOS, and H IF-2α/VEGF to antagonize Hx-induced HCMEC injury.

Effect of Serum from Overfatigue Rats on JNK/c-Jun/HO-1 Pathway in Human Umbilical Vein Endothelial Cells and the Intervening Effect of Tongxinluo(通心络)Superfine Powder

Objective:To cultivate human umbilical vein endothelial cells (HUVECs) in the serum of overfatigue rats with the intervention of Tongxinluo (通心络) superfine powder (TXLSP).By examining the variation of the activity of JNK/c-Jun/HO-1 pathway,the possible mechanisms of vascular endothelial dysfunction under overfatigue conditions and the intervening effect of TXLSP were explored.Methods:The HUVECs were randomly divided into the normal control group,the model group,the SP600125 (a specific antagonist of JNK)gro...

Effects of Chinese medicine Tongxinluo on hyperglycemia and beta-cell damage in streptozotocin-induced diabetic rats

Background Oxidative stress has been implicated in the onset and progression of diabetes. Tongxinluo is a traditional Chinese medicine with potent antioxidant properties. The aim of this study was to test the hypothesis that pretreatment with Tongxinluo has similar effects as melatonin on preventing hyperglycemia and beta-cell damage in a rat model of streptozotocin （STZ）-induced diabetes. Methods Forty male Sprague Dawley rats were randomly assigned to four groups （n=10 each）： normal control （NC） group; STZ group （70 mg/kg, i.p.）; Tongxinluo （1.0 g-kg-^-d-1） pretreated （TXL＋STZ） group and melatonin （200 iJg-kg-~.d-1） pretreated （MLT＋STZ） group. Tongxinluo and melatonin were administered by gavage beginning 8 days before STZ injection and continuing until the end of the study （15 days after STZ administration）. Blood glucose levels and body weights, malondialdehyde （MDA）, and reduced glutathione （GSH） levels were measured, and immunofluorescence studies were performed in all of the groups. Results Pretreatment with Tongxinluo, as with melatonin, attenuated severe hyperglycemia and weight loss induced by STZ.. In pancreatic homogenates, MDA levels were significantly lower and GSH levels were significantly higher in Tongxinluo pretreated group and in melatonin pretreated group than those in STZ group. Values of insulin staining were significantly improved in Tongxinluo pretreated group and in melatonin pretreated group as compared with those in STZ group. Conclusions Tongxinluo, as melatonin, prevented hyperglycemia and beta-cell destruction induced by STZ in rats through reducing oxidative stress in pancreatic tissues. Tongxinluo may provide an alternative therapy for the prevention and treatment of diabetes.

Tongxinluo Improves Apolipoprotein E-Deficient Mouse Heart Function

Background： Our previous studies have shown that Tongxinluo （TXL）, a compound Chinese medicine, can decrease myocardial ischemia-reperfusion injury, protect capillary endothelium function, and lessen cardiac ventricle reconstitution in animal models. The aim of this study was to illuminate whether TXL can improve hypercholesterolemia-impaired heart function by protecting artery endothelial function and increasing microvascular density （MVD） in heart. Furthermore, we will explore the underlying molecular mechanism of TXL cardiovascular protection. Methods： After intragastric administration of TXL （0.1 ml/10 g body weight） to C57BL/6J wild-type mice （n = 8） and ApoE-/- mice （n=8）, total cholesterol, high-density lipoprotein-cholesterol, very-low-density lipoprotein （VLDL）-cholesterol, triglyceride, and blood glucose levels in serum were measured. The parameters of heart rate （HR）, lelt ventricular diastolic end diameter, and left ventricular systolic end diameter were harvested by ultrasonic cardiogram. The left ventricular ejection fraction, stroke volume, cardiac output, and let1 ventricular fractional shortening were calculated. Meanwhile, aorta peak systolic flow velocity （PSV）, end diastolic flow velocity, and mean flow velocity （MFV） were measured. The pulsatility index （PI） and resistant index were calculated in order to evaluate the vascular elasticity and resistance. Tile endothelium-dependent vasodilatation was evaluated by relaxation of aortic rings in response to acetylcholine. Western blotting and real-time quantitative reverse transcription polymerase chain reaction were performed for protein and gene analyses of vascular endothelial growth factor （VEGF）. lnamunohistochenaical detection was perlbrmed for myocardial CD34 expression. Data in this study were compared by one-way analysis of variance between groups. A value of P 〈 0.05 was considered statistically significant. Results： Although there was no significant decrease of cholesterol level （F = 2.300, P = 0.240）, TXL inhibited the level of triglyceride and VLDL （F- 9.209, P- 0.024 and F = 9.786, P ： 0.020, respectively） in ApoE-/- mice. TXL improved heart function of ApoE-/- mice owing to the elevations of LVEF, SV, CO, and LVFS （all P 〈 0.05）. TXL enhanced aortic PSV and MFV （F = 10.774, P = 0.024 and F = 11.354, P - 0.020, respectively） and reduced PI of ApoE-/- mice （1.41 ± 0.17 vs. 1.60 ± 0.17; P= 0.037）. After incubation with 10μmol/L acetylcholine, the ApoE-/- mice treated with TXL aortic segment relaxed by 44% - 3%, significantly higher than control group mice （F = 9.280, P = 0.040）. TXL also restrain the angiogenesis of ApoE-/- mice aorta （F = 21.223, P = 0.010）. Compared with C57BL/6J mice, the MVD was decreased in heart tissue of untreated ApoE-/- mice （54.0 ± 3.0/mm2 vs. 75.0 ± 2.0/mm2; F = 16.054, P - 0.010）. However, TXL could significantly enhance MVD （65.0 ± 5.0/mm2 vs. 54.0 ±3.0/mm2; F- 11.929, P = 0.020） in treated ApoE-/- mice. In addition, TXL obviously increased the expression of VEGF protein determined by Western blot （F = 20.247, P = 0.004）. Conclusions： TXL obviously improves the ApoE-/- mouse heart function from different pathways, including reduces blood fat tolessen atherosclerosis; enhances aortic impulsivity, blood supply capacity, and vessel elasticity： improves endothelium-dependent vasodilatation： restraines angiogenesis of aorta-contained plaque; and enhances MVD of heart. The molecular mechanism of MVD enhancement maybe relate with increased VEGF expression.

Observation of changes in the number of myocardial capillaries in rabbits after treatment of acute myocardial infarction by Tongxinluo superfine powder

OBJECTIVE: To investigate the effects of Tongxinluo superfine powder on cardiac function, infarct size and the number of myocardial capillaries in a rabbit model of acute myocardial infarction.METHODS: A total of 32 New Zealand white rabbits were randomly divided into four groups: sham operation group, model group, treatment group,and pre-treatment, the experiment of pre-treatment group was performed 6 weeks early than the treat) group,The four groups use a unified modeling technique. An acute myocardial infarction model was established through external application of70% ferric chloride on the coronary artery. After 7 d,electrocardiogram, ultrasonography of cardiac function, micro-computed tomography, pathology and other data were collected.RESULTS: In the treatment and pre-treatment groups, ejection fraction, left ventricular short axis shortening rate, left ventricular end-systolic diameter and cardiac output significantly improved, the number of capillaries significantly increased, and infarct size significantly decreased. In addition, the results suggest that the value of intra-ventricular pressure and the situation of electrocardiogram also changed to different degrees with the increasing of treatment of cycle.CONCLUSION: Tongxinluo superfine powder can protect the myocardium, improve the blood supply of the myocardium and reduce the degree of myocardial injury, during acute stage of myocardial infarction.

Tongxinluo promotes mesenchymal stem cell tube formation in vitro

Objective： To study whether Tongxinluo （TXL） can induce angiogenesis in bone marrow mesenchymal stem cells （MSCs）, and to investigate the underlying mechanism. Methods： Bone marrow MSCs were obtained from male Sprague-Dawley rats. We established an angiogenesis model in vitro via matrigel experiment. MSCs were seeded on matrigel coated 24-well plates, and treated by TXL 50 and 100 mg/L. After 24 h, we observed the tube formations of MSCs in the matrigel. Cell migration ability was examined by wound scratch test and transwell assay. Expressions of vascular endothelial growth factor （VEGF）, fetal liver kinase-1 （FIk-1）, matrix metalloproteinase-2 （MMP-2）, MMP-9, and tissue inhibitor of metalloproteinase-2 （TIMP-2） were analyzed at the protein level by Western blot. Gelatin zymography assay was applied to investigating the MSC paracrine abilities of pro-MMP-2 and activated MMP-2. Results： TXL promoted MSC tube formation in matrigel. The ratio of TXL 100 mg/L treated-MSC tubular length was increased 3.04-fold compared to the control group （P〈0.05）. Scratch test and transwell assay showed that TXL could improve the cell migration ability of MSCs. Western blot experiments showed that TXL promoted MSC synthesis of MMP-2, but it had no influence on the expressions of MMP-9 and TIMP-2. This effect was confirmed by gelatin zymography assay, which showed that TXL increased MSC secretion of pro-MMP-2 and activated MMP-2. VEGF expression of TXL treated-MSCs was increased compared to the control group. The expression of FIk-1 was not different among the groups. Conclusions： This study demonstrates that TXL can promote the tube formation of MSCs, and the underlying mechanisms are associated with increased migration ability of MSCs and the up-regulation of MMP-2 and VEGF expressions.

Protective Effects and Potential Mechanism of Tongxinluo on Mice with Thromboangiitis Obliterans Induced by Sodium Laurate

Objective To investigate the effects of Tongxinluo(TXL)on thromboangiitis obliterans(TAO)and the underlying mechanisms.Methods Ninety male C57/BL6J mice were randomly divided into 6 groups according to a random number table:the sham group,TAO model group,Compound Danshen Tablet(CDT)group,and the high-,medium-,and low-dose TXL groups.All mice except the sham group were injected with sodium laurate(0.1 mL,5 mg/mL)in the femoral artery to establish TAO mouse model.After modeling,mice in the sham and TAO model groups were intragastrically administered 0.5%(w/v)sodium carboxymethylcellulose,mice in the CDT group were intragastrically administered 0.52 g/kg CDT,and mice in the TXL-H,TXL-M,and TXL-L groups were intragastrically administered 1.5,0.75,and 0.38 g/kg TXL,respectively.After 4 weeks of gavage,the recovery of blood flow in the lower limbs of mice was detected by Laser Doppler Imaging.The pathological changes and thrombosis of the femoral artery were observed by morphological examination.The expressions of tumor necrosis factorα(TNF-α)and inducible nitric oxide synthase(iNOS)in the femoral artery wall were detected by HE staining.Levels of thromboxane B2(TXB2),6-keto-prostaglandin F1α(6-keto-PGF1α),endothelin-1(ET-1),interleukin(IL)-1βand IL-6 were measured using enzyme-linked immunosorbent assay(ELISA).Levels of activated partial thromboplastin time(APTT),prothrombin time(PT),thrombin time(TT)and fibrinogen(FIB)were detected by a fully automated biochemical analyzer.Results TXL promoted the restoration of blood flow in the lower limbs,reduced the area of thrombosis in the femoral artery,and alleviated the pathological changes in the femoral artery wall.Moreover,the levels of TXB2,ET-1,IL-6,IL-1β,TNF-αand iNOS were significantly lower in the TXL groups compared with the model group(P<0.05 or P<0.01),while the level of 6-keto-PGF1αwas significantly higher(P<0.01).In addition,APTT,PT,and TT were significantly prolonged in TXL groups compared with the model group(P<0.05 or P<0.01),and FIB levels were significantly decreased compared with the model group(P<0.01).Conclusions TXL had a protective effect on TAO mice,and the mechanism may involve inhibition of thrombosis and inflammatory responses.TXL may be a potential drug for the treatment of TAO.

Early Intervention of Tongxinluo(通心络) on Right Ventricular Function Assessed by Echocardiography in Rats with Pulmonary Arterial Hypertension Induced by Monocrotaline

Objective:To investigate the effect of early intervention of Tongxinluo(通心络,TXL)on right ventricular function(RVF)of rats with pulmonary arterial hypertension(PAH)induced by monocrotaline(MCT).Methods:A total of 30 adult male Sprague-Dawley rats were assigned to 5 groups with complete random experiment design:Sham group(Sham),MCT group,TXL group,sildenafil(SIL)group and combination group(TXL+SIL),6 rats in each group.Rats were injected with 50 mg/kg MCT solution for inducing PAH model except for those in the sham group.From the day of modeling,rats of TXL,SIL and TXL+SIL groups were given TXL(1.2 g/kg),SIL(10 mg/kg)and combination solution(TXL:1.2 g/kg,SIL:10 mg/kg)respectively,and rats in Sham and MCT groups were given normal saline(5 m L/kg).The samples were collected and tested after 21 consecutive days of intragastric administration.Echocardiography was used to measure the related indices of RVF,including pulmonary arterial flow spectrum,pulmonary artery diameter(PAD),right ventricular wall thickness(RVWT),right ventricular diameter(RVD),tricuspidannular plane systolic excursion(TAPSE),right atrium transverse diameter(RAT),and inferior vena cava diameter(IVCD).Elastic Verhoeff-Van Gieson staining was adopted to measure the percentage of wall thickness(WT%)of pulmonary arteriols.Hematoxylin-eosin staining was used to measure the cross-sectional area(CSA)of right ventricular cardiomyocytes.Results:MCT-induced PAH rat model was successfully established.In MCT group the wall of pulmonary arterioles exhibited a prominent-increase thickness,PAD,RVWT,RVD,RAT,IVCD,WT%,right ventricular hypertrophy index(RVHI)as well as CSA of RV cardiomyocyte significantly increased(all P<0.01),and TAPSE markedly decreased(P<0.01).At the same time,TXL prominently improved all of the above indices(all P<0.01).In comparison with SIL,TXL significantly reduced RVD(P<0.05)and decreased CAS of RV cardiomyocytes(P<0.01),but TAPSE in SIL group was much larger than in TXL group(P<0.01).Moreover,TAPSE in TXL+SIL group was larger than that in TXL group(P<0.01),while the two groups performed equally well in terms of the other indices.Conclusion:Early intervention of TXL could inhibit pulmonary arterioles remodeling,and improve RVF by attenuating right ventricular hypertrophy,and TXL has a stronger effect on inhibiting right ventricular remodeling than SIL.

通心络防治冠心病临床应用中国专家共识

《通心络防治冠心病临床应用中国专家共识》严格遵循共识制订规范,以循证为主、共识为辅、经验为鉴,旨在规范通心络在治疗冠心病中的合理应用。针对常见类型的冠心病,在常规治疗基础上联用通心络可增加临床获益。

通心络胶囊治疗脓毒症疗效及对微循环的影响

目的观察通心络胶囊对脓毒症患者临床疗效和微循环指标的影响。方法选取2018年11月—2021年10月河北以岭医院ICU收治的脓毒症患者60例,按照随机数字表法分为观察组和对照组,各30例。2组均给予西医常规治疗,观察组加用通心络胶囊,疗程均7 d。分别于入ICU 0、6、24、48、72 h检测大循环参数(MAP、CVP及ScvO_(2))、微循环参数(Pcv-aCO_(2)、Pcv-aCO_(2)/Ca-cvO_(2)、Lac、CRT)及APACHE-Ⅱ、SOFA评分、机械通气时间、ICU住院时间及28 d病死率。结果观察组临床疗效总有效率显著高于对照组(100.0%vs.66.7%,χ^(2)=8.857,P=0.003);2组0、6、24、48及72 h不同时间点大循环参数MAP、CVP及ScvO_(2)组内比较均具有统计意义(P均<0.001);而组间比较皆无统计意义(P均>0.05)。对照组0、6、24、48及72 h不同时间点Pcv-aCO_(2)组内比较差异具有统计学意义(P=0.006),Pcv-aCO_(2)/Ca-cvO_(2)组内比较无统计学意义(P=0.261);观察组Pcv-aCO_(2)及Pcv-aCO_(2)/Ca-cvO_(2)组内比较差异均具有统计学意义(P均<0.05);2组Pcv-aCO_(2)及Pcv-aCO_(2)/Ca-cvO_(2)在0、6、24 h组间比较差异均无统计学意义(P>0.05),但48及72 h组间比较均具有统计学意义(P均<0.05),观察组均优于对照组。2组0、6、24、48及72 h不同时间点Lac、CRT组内比较均具有统计学意义(P均<0.001);2组0、6、24、48 h Lac组间比较均无统计学意义(P均>0.05),72 h Lac组间比较差异具有统计学意义(P=0.036),观察组优于对照组;2组0、6、24 h CRT组间比较均无统计学意义(P均>0.05),48及72 h CRT组间比较差异均具有统计学意义(P均<0.05),观察组均优于对照组。2组7 d与1 d组内比较APACHE-Ⅱ及SOFA评分差异均有统计学意义(P均<0.001);2组APACHE-Ⅱ与SOFA评分1 d组间比较差异均无统计学意义(P均>0.05);7 d时APACHE-Ⅱ评分组间比较差异无统计学意义(P>0.05),SOFA评分组间比较差异有统计学意义(P=0.010),观察组优于对照组。2组在机械通气时间、ICU住院时间及28 d病死率方面比较差异均无统计学意义(P均>0.05)。结论通心络胶囊能够改善脓毒症患者微循环障碍,缓解病情,改善中医证候临床疗效,但未能缩短机械通气时间、ICU住院时间及28 d病死率。

通心络对高糖高脂诱导人主动脉内皮细胞损伤的保护作用

目的探讨高糖高脂对人主动脉内皮细胞(HAECs)的影响及通心络的干预作用。方法经随机方式,将HAECs分为以下5组:通心络高、中、低浓度组,高糖高脂组和空白对照组。检测各组细胞生存活性、细胞上清液乳酸脱氢酶(LDH)含量、三磷酸腺苷(ATP)含量、线粒体膜电位(MMP)、细胞内活性氧簇(ROS)水平和细胞色素(Cyt)-C蛋白表达。结果与空白对照组比较,高糖高脂组MMP、ATP水平与细胞生存活性明显下降,细胞中ROS含量、细胞上清液LDH漏出量与Cyt-C蛋白表达皆明显上升(P<0.05,P<0.01);与高糖高脂组相比,通心络干预后细胞生存活性、ATP含量和MMP明显升高,细胞上清液LDH漏出量、细胞内ROS水平和Cyt-C蛋白表达明显降低(P<0.05,P<0.01)。结论对于高糖高脂损伤的HAECs,通心络具有保护作用,相关保护机制可能和保护线粒体功能相关。